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Title: Crystallization and preliminary X-ray crystallographic analysis of EstE1, a new and thermostable esterase cloned from a metagenomic library

Abstract

Recombinant EstE1 protein with a histidine tag at the C-terminus was overexpressed in Escherichia coli strain BL21(DE3) and then purified by affinity chromatography. The protein was then crystallized at 290 K by the hanging-drop vapour-diffusion method. EstE1, a new thermostable esterase, was isolated by functional screening of a metagenomic DNA library from thermal environment samples. This enzyme showed activity towards short-chain acyl derivatives of length C4–C6 at a temperature of 303–363 K and displayed a high thermostability above 353 K. EstE1 has 64 and 57% amino-acid sequence similarity to est{sub pc}-encoded carboxylesterase from Pyrobaculum calidifontis and AFEST from Archaeoglobus fulgidus, respectively. The recombinant protein with a histidine tag at the C-terminus was overexpressed in Escherichia coli strain BL21(DE3) and then purified by affinity chromatography. The protein was crystallized at 290 K by the hanging-drop vapour-diffusion method. X-ray diffraction data were collected to 2.3 Å resolution from an EstE1 crystal; the crystal belongs to space group P4{sub 1}2{sub 1}2, with unit-cell parameters a = b = 73.71, c = 234.23 Å. Assuming the presence of four molecules in the asymmetric unit, the Matthews coefficient V{sub M} is calculated to be 2.2 Å{sup 3} Da{sup −1} and the solvent content is 44.1%.

Authors:
 [1];  [2];  [3];  [1];  [3];  [1];  [2]
  1. Department of Biology, Yonsei University, Seoul 120-749 (Korea, Republic of)
  2. (Korea, Republic of)
  3. Department of Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of)
Publication Date:
OSTI Identifier:
22356264
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section F; Journal Volume: 62; Journal Issue: Pt 2; Other Information: PMCID: PMC2150951; PMID: 16511287; PUBLISHER-ID: fw5062; OAI: oai:pubmedcentral.nih.gov:2150951; Copyright (c) International Union of Crystallography 2006; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United Kingdom
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; AFFINITY; CRYSTALLIZATION; CRYSTALS; DIFFUSION; ENVIRONMENT; ESCHERICHIA COLI; HISTIDINE; IRON; MOLECULES; RESOLUTION; SCREENING; SPACE GROUPS; STRAINS; X-RAY DIFFRACTION

Citation Formats

Byun, Jung-Sue, Protein Network Research Center, Yonsei University, Seoul 120-749, Rhee, Jin-Kyu, Kim, Dong-Uk, Oh, Jong-Won, Cho, Hyun-Soo, E-mail: hscho8@yonsei.ac.kr, and Protein Network Research Center, Yonsei University, Seoul 120-749. Crystallization and preliminary X-ray crystallographic analysis of EstE1, a new and thermostable esterase cloned from a metagenomic library. United Kingdom: N. p., 2006. Web. doi:10.1107/S1744309106000832.
Byun, Jung-Sue, Protein Network Research Center, Yonsei University, Seoul 120-749, Rhee, Jin-Kyu, Kim, Dong-Uk, Oh, Jong-Won, Cho, Hyun-Soo, E-mail: hscho8@yonsei.ac.kr, & Protein Network Research Center, Yonsei University, Seoul 120-749. Crystallization and preliminary X-ray crystallographic analysis of EstE1, a new and thermostable esterase cloned from a metagenomic library. United Kingdom. doi:10.1107/S1744309106000832.
Byun, Jung-Sue, Protein Network Research Center, Yonsei University, Seoul 120-749, Rhee, Jin-Kyu, Kim, Dong-Uk, Oh, Jong-Won, Cho, Hyun-Soo, E-mail: hscho8@yonsei.ac.kr, and Protein Network Research Center, Yonsei University, Seoul 120-749. Wed . "Crystallization and preliminary X-ray crystallographic analysis of EstE1, a new and thermostable esterase cloned from a metagenomic library". United Kingdom. doi:10.1107/S1744309106000832.
@article{osti_22356264,
title = {Crystallization and preliminary X-ray crystallographic analysis of EstE1, a new and thermostable esterase cloned from a metagenomic library},
author = {Byun, Jung-Sue and Protein Network Research Center, Yonsei University, Seoul 120-749 and Rhee, Jin-Kyu and Kim, Dong-Uk and Oh, Jong-Won and Cho, Hyun-Soo, E-mail: hscho8@yonsei.ac.kr and Protein Network Research Center, Yonsei University, Seoul 120-749},
abstractNote = {Recombinant EstE1 protein with a histidine tag at the C-terminus was overexpressed in Escherichia coli strain BL21(DE3) and then purified by affinity chromatography. The protein was then crystallized at 290 K by the hanging-drop vapour-diffusion method. EstE1, a new thermostable esterase, was isolated by functional screening of a metagenomic DNA library from thermal environment samples. This enzyme showed activity towards short-chain acyl derivatives of length C4–C6 at a temperature of 303–363 K and displayed a high thermostability above 353 K. EstE1 has 64 and 57% amino-acid sequence similarity to est{sub pc}-encoded carboxylesterase from Pyrobaculum calidifontis and AFEST from Archaeoglobus fulgidus, respectively. The recombinant protein with a histidine tag at the C-terminus was overexpressed in Escherichia coli strain BL21(DE3) and then purified by affinity chromatography. The protein was crystallized at 290 K by the hanging-drop vapour-diffusion method. X-ray diffraction data were collected to 2.3 Å resolution from an EstE1 crystal; the crystal belongs to space group P4{sub 1}2{sub 1}2, with unit-cell parameters a = b = 73.71, c = 234.23 Å. Assuming the presence of four molecules in the asymmetric unit, the Matthews coefficient V{sub M} is calculated to be 2.2 Å{sup 3} Da{sup −1} and the solvent content is 44.1%.},
doi = {10.1107/S1744309106000832},
journal = {Acta Crystallographica. Section F},
number = Pt 2,
volume = 62,
place = {United Kingdom},
year = {Wed Feb 01 00:00:00 EST 2006},
month = {Wed Feb 01 00:00:00 EST 2006}
}
  • The crystallization of ybfF, a new esterase from E. coli, and the collection of diffraction data to 1.1 Å resolution are reported. The product of the recently discovered ybfF gene, which belongs to the esterase family, does not show high sequence similarity to other esterases. To provide the molecular background to the enzymatic mechanism of the ybfF esterase, the ybfF protein from Escherichia coli K12 (Ec-ybfF) was cloned, expressed and purified. The Ec-ybfF protein was crystallized from 60% Tacsimate and 0.1 M bis-Tris propane buffer pH 7.0. Diffraction data were collected to 1.10 Å resolution using synchrotron radiation. The crystalmore » belongs to the orthorhombic space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 66.09, b = 90.71, c = 92.88 Å. With two Ec-ybfF molecules in the asymmetric unit, the crystal volume per unit protein weight is 2.17 Å{sup 3} Da{sup −1}, corresponding to a solvent content of 42%.« less
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  • The catalytic domain of the glucuronoyl esterase from H. jecorina was overexpresssed, purified and crystallized in space group P2{sub 1}2{sub 1}2{sub 1}. X-ray diffraction data were collected to 1.9 Å resolution. The catalytic domain of the glucuronoyl esterase from Hypocrea jecorina (anamorph Trichoderma reesei) was overexpresssed, purified and crystallized by the sitting-drop vapor-diffusion method using 1.4 M sodium/potassium phosphate pH 6.9. The crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1} and X-ray diffraction data were collected to 1.9 Å resolution. This is the first enzyme with glucoronoyl esterase activity to be crystallized; its structure will be valuable in lignocellulose-degradationmore » research.« less